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Biomedical subjects

B Palcic

Publications and source records attributed to B Palcic.

At least 19 recordsLinked to original sources

Automated image detection and segmentation in blood smears.

A simple technique which automatically detects and then segments nucleated cells in Wright's giemsa-stained blood smears is presented. Our method differs from others in 1) the simplicity of our algorithms; 2) inclusion of touching (as well as nontouching) cells; and 3) use of these algorithms to segment as well as to detect nucleated cells employing conventionally prepared smears. Our method involves: 1) acquisition of spectral images; 2) preprocessing the acquired images; 3) detection of single and touching cells in the scene; 4) segmentation of the cells into nuclear and cytoplasmic regions; and 5) postprocessing of the segmented regions. The first two steps of this algorithm are employed to obtain high-quality images, to remove random noise, and to correct aberration and shading effects. Spectral information of the image is used in step 3 to segment the nucleated cells from the rest of the scene. Using the initial cell masks, nucleated cells which are just touching are detected and separated. Simple features are then extracted and conditions applied such that single nucleated cells are finally selected. In step 4, the intensity variations of the cells are then used to segment the nucleus from the cytoplasm. The success rate in segmenting the nucleated cells is between 81 and 93%. The major errors in segmentation of the nucleus and the cytoplasm in the recognized nucleated cells are 3.5% and 2.2%, respectively.

Algorithms

Detection of squamous cell cancer and pre-cancerous lesions by imaging of tissue autofluorescence in the hamster cheek pouch model.

Early detection of invasive and pre-invasive neoplasms of the aerodigestive tract will ultimately improve the management of patients with these lesions. This paper describes the use of quantitative fluorescence imaging of early squamous cell carcinomas in an animal model. Dysplasia, carcinoma in situ and invasive cancers were imaged exploiting tumour autofluorescence. Mapped biopsies were obtained from areas imaged determining a sensitivity of 100% and specificity of 80%. Autofluorescence imaging is an excellent method of detecting neoplasms of the aerodigestive tract.

9,10-Dimethyl-1,2-benzanthracene

The relative biological effectiveness of 60Co gamma-rays, 55 kVp X-rays, 250 kVp X-rays, and 11 MeV electrons at low doses.

The relative biological effectiveness (RBE) of selected low-LET radiation modalities (55 kVp X-rays, 250 kVp X-rays, 60Co gamma-rays, and 11 MeV electrons) was investigated for survival of two cell lines (V79 and CHO). Detailed measurements were made in the low (0 to 3 Gy) dose range using an image cytometry device to accurately determine the number of cells assayed at each dose point. Data were also collected in the high dose range (0 to 10 Gy) using conventional counting and plating techniques. RBE values (+/- 1 SE) varied from 1.0 +/- 0.07 (V79 cells) and 1.2 +/- 0.05 (CHO cells) at high doses to 1.3 +/- 0.07 (V79) and 1.4 +/- 0.1 (CHO) at low doses for 55 kVp X-rays, from 1.1 +/- 0.05 (V79) and 1.1 +/- 0.04 (CHO) at high doses to 1.1 +/- 0.06 (V79) and 1.2 +/- 0.2 (CHO) at low doses for 250 kVp X-rays, and from 1.1 +/- 0.08 (V79) and 1.0 +/- 0.04 (CHO) at high doses to 1.0 +/- 0.06 (V79) and 0.9 +/- 0.1 (CHO) at low doses for 11 MeV electrons. Only the low and high dose RBEs for 55 kVp X-rays relative to 60Co gamma-rays were significantly different.

Animals

Detection of dysplasia and carcinoma in situ by ratio fluorometry.

Fluorescence bronchoscopy was performed in 82 volunteers recruited from occupational groups at risk of exposure to asbestos and/or diesel fumes to determine whether differences in tissue autofluorescence between normal and malignant bronchial tissues can be used to improve the sensitivity of standard fiberoptic bronchoscopy in detecting dysplasic and carcinoma in situ (CIS). This study consisted of 25 nonsmokers, 40 exsmokers, and 17 current smokers with mean ages of 52, 55, and 49 yr, respectively. Tissue autofluorescence was induced by a blue light from an He-Cd laser coupled to the illumination channel of the bronchoscope and analyzed by a ratiofluorometer. One or more sites of moderate or severe dysplasia were found in 12% of the exsmokers and current smokers but in none of the nonsmoker volunteers. CIS was found in two of the exsmokers. The sensitivity of fluorescence bronchoscopy (86%) was found to be 50% better than that of conventional white-light bronchoscopy (52%) in detecting dysplasia and CIS. Pre- and post-bronchoscopy sputum cytology failed to detect these precancerous lesions. Our results suggest that fluorescence bronchoscopy may be an important new method that can improve the ability to detect and localize precancerous and/or CIS lesions.

Asbestos

Comparison of three different methods for automated classification of cervical cells.

Over 4600 exfoliated squamous cervical cells taken from appropriate Papanicolaou samples were classified as normal, mildly dysplastic, moderately dysplastic and severely dysplastic by an experienced cytopathologist. The slides were de-stained and subsequently re-stained with Feulgen Thionin-SO2 stain. Images of the nuclei were then captured, recorded and processed employing an image cytometry device. Automated classification of the cells was carried out using three different methods--discriminant function analysis, a decision tree classifier and a neutral network classifier. The discriminant function analysis method, which combined all dysplastic cells into an abnormal group, achieved a combined error rate of less than 0.4% for moderate and severe dysplastic cells, and less than 40% for mildly dysplastic cells. All three methods yielded comparable results, which approached those of human performance.

Cell Nucleus

Determination of DNA ploidy in archival tissue from non-Hodgkin's lymphoma using flow and image cytometry.

Paraffin-embedded tissue from a series of 40 cases of diffuse, large cell lymphoma was analyzed by both flow and image cytometry to compare the ability of these techniques to detect DNA aneuploid populations. Image cytometry (ICM) was performed both on nuclear suspensions and tissue sections. Twenty cases (50%) were non-diploid by at least one method of analysis. Twenty-five percent of the cases were aneuploid by flow cytometry (FCM) alone. The majority of these cases were near-diploid tumors which could not be resolved by ICM. Peri-tetraploid peaks were identified by ICM of tissue sections alone in 15% of the cases. There was an apparent loss of these peri-tetraploid cells during the preparation of the nuclear suspensions. The remaining cases showed a good correlation between all three methods in the determination of DNA ploidy. Flow and image cytometry are complimentary techniques when applied to archival tissue, however aneuploid populations may be missed if ICM is not performed on tissue sections.

Aneuploidy

Autofluorescence of normal and malignant bronchial tissue.

In vivo autofluorescence spectra were obtained in 5 patients with carcinoma in situ, 26 patients with invasive tumors, and 1 patient with severe dysplasia. Significant spectral differences were observed between pre-cancerous, cancerous, and normal bronchial tissues. This difference may afford a method to image and/or detect early lung cancer by using tissue autofluorescence alone.

Bronchi

Computer automation in measurement and analysis of cell motility in vitro.

The application of computerized techniques to the study of cell motility has allowed direct examination and quantitation of individual cell movement and morphology. This has stimulated interest in theoretical models of cell motility, and has prompted the need for increasingly sophisticated approaches to analyze and present cell motility data. In this article we review some of the microscope systems currently used to study cell motility, and give more details on such a system developed in our own laboratory, the DMIPS Cell Analyzer. Brief examples of its application to the field of cell motility are provided. Some areas of further research in the field of automated microscopy are also discussed.

Animals

Effect of focus on cell detection and recognition by the Cell Analyzer.

The effect of defocusing on the quality of signals from live cells detected by an automated image cytometry device, the Cell Analyzer, was examined. The influence of these effects on the ability of this device to automatically locate cells plated into a tissue culture flask was then determined by measuring the performance of cell detection and recognition procedures as a function of focus setting. Acceptable limits for deviation from the optimal focus setting (as determined by microscope objective position) were found to be similar for both these procedures, ranging from 40 microns below to 25 microns above the optimal focus position. These limits were asymmetrical about ideal focus due to a pronounced asymmetry in the effects of positive and negative defocusing on the cell signal.

Animals

The effect of retinoids, carotenoids and phenolics on chromosomal instability of bovine papillomavirus DNA-carrying cells.

Antioxidants were found to protect against the genotoxic effects of chemical and physical mutagenic and clastogenic agents. This study focused on the capacity of antioxidants to reduce an intrinsic and persistent chromosome instability. As a model system, strains of C127 cells, which were transformed by bovine papillomavirus (BPV) DNA and which carry BPV DNA varying from 20 to 160 copies, were used. Transformed cells of 10 different strains showed a persistently high incidence of mitotic irregularities detectable at anaphase and telophase (27.3-58.9%), an elevated frequency of cells with micronuclei (6.6-34.7%), and a broad spectrum of nuclear sizes, as measured by image analysis. A 3-day exposure to retinoic acid, retinol, beta-carotene, canthaxanthin, ascorbic acid and ellagic acid greatly reduced the degree of chromosome instability, whereas catechin, eugenol and pyrogallol showed a smaller inhibitory effect, and curcumin had no detectable effect on the frequency of mitotic irregularities. After withdrawal of retinoic acid treatment, the high levels of chromosome instability reappeared. The possibility that the protective effect of the retinoids and carotenoids examined in the model system points to their beneficial administration to human cells with an intrinsic or acquired chromosome instability is discussed.

Antioxidants

The effects of low density lipoproteins on uptake of Photofrin II.

The cellular uptake of Photofrin II (PII) was studied using fluorescence imaging and chemical extraction. The influence of serum and low density lipoproteins (LDL) was examined under a variety of experimental conditions employing cultured human cells of different origins as well as a subcutaneously SMT-F tumor implanted in mice. Results showed that serum inhibited PII uptake. In general, LDL also inhibits PII uptake with the exception of an initial increase in the first 10-30 min when the cellular concentration of PII was measured by fluorescence imaging instead of chemical extraction. Our results suggest a possible de-aggregation process occurring upon internalization or binding of PII to LDL.

Animals

Detection of early lung cancer using low dose Photofrin II.

Fluorescence imaging using hematoporphyrin derivative (HpD) or Photofrin II as a tumor marker has been used for localization of early bronchogenic carcinoma. Wider clinical application of HpD or Photofrin II as a cancer imaging agent has been hampered by the potentially serious and prolonged skin photosensitivity. Using a sensitive fluorescence bronchoscope system with a ratio fluorometer probe, carcinoma in situ was detected in four patients with low dose Photofrin II (0.25 mg/kg) with no apparent skin phototoxicity to 30 J/cm2 visible light on skin photosensitivity test.

Aged

Fractal texture features based on optical density surface area. Use in image analysis of cervical cells.

The spatial distribution of the optical density of an image of a cell nucleus can be imagined to resemble a rugged surface in a three-dimensional space. The area of this surface can be measured using different unit-sized squares, yielding different values for the total area depending on the unit size. A fractal dimension can be calculated from the relationship between the unit size and the overall area measured; this fractal dimension is characteristic of the surface measured. This technique was applied to extract three new nuclear features from stained cervical cells. These new features were useful for the computer discrimination of normal and premalignant cervical cells. The features are defined, and their discriminating power is compared to that of some other nuclear features.

Cell Membrane

Quantitation of chromatin patterns by image analysis as a predictive tool in chemopreventive trials with vitamin A.

Nuclear textures of oral mucosal cells were quantitated by image analysis, and their suitability as markers in a chemopreventive trial explored. Subjects were chewers of tobacco-containing betel quids with well established oral leukoplakias. Treatment consisted of a weekly oral administration of vitamin A (200,000 IU/week) for six months. Leukoplakias regressed in 57.1% of the 21 trial participants. The original leukoplakias did not redevelop within four months after termination of treatment. For image analysis, biopsies were taken from leukoplakias of five chewers before administration of vitamin A, at the end of the administration, and four months after termination of treatment. Sections of paraffin-embedded biopsies were stained with the Feulgen reaction and submitted to quantitative image analysis of two parameters: variance of intensity and entropy. Both these parameters were significantly reduced in all five trial participants as a result of the six-month vitamin A treatment. During the post-treatment period, nuclei with condensed chromatin, as measured by the variance of intensity, reappeared in the mucosa of four of the five chewers examined, although no leukoplakias were detectable on visual examination of the oral cavity. The results indicate that the quantitation of nuclear textures in a small subpopulation of a chemopreventive trial could conceivably be a simple marker with a predictive value.

Biomarkers, Tumor

Pharmacological modulation of human subconjunctival fibroblast behavior in vitro.

The response of human subconjunctival fibroblasts to a variety of pharmacological agents was evaluated utilizing a novel in vitro wound assay and a separate proliferation assay. Both colchicine and cytochalasin B dramatically arrested wound closure at concentrations greater than or equal to 0.01 micrograms/ml and 2 micrograms/ml, respectively (p less than 0.05). At lower doses these drugs altered fibroblast morphology and inhibited directed cell migration. Dexamethasone and 6-MP delayed wound closure at concentrations greater than or equal to 100 micrograms/ml and 1000 micrograms/ml, respectively (p less than 0.05). Effective antiproliferative agents, in order of decreasing potency (based on unit weight), were Cytarabine (cytosine arabinoside), doxorubicin (Adriamycin), colchicine, 5-fluorouracil, cytochalasin B, cyclosporin (Sandimmune), 6-mercaptopurine, and dexamethasone. The antiprotease agents and methotrexate were ineffective as determined by both assays. We conclude that the wound assay is well suited for rapid screening of drugs for their effect on fibroblast morphology, motility, and proliferation, and that colchicine and cytochalasin B, in doses well below those documented to produce ocular toxicity, are effective in inhibiting directed migration and proliferation of subconjunctival fibroblasts in vitro. Differences in mechanism, onset of action, therapeutic range, and cytotoxicity of drugs could be exploited in controlling ocular fibroblast behavior in vivo.

Antineoplastic Agents

An edge relocation segmentation algorithm.

An automated procedure that refines the nuclear contour of a previously segmented nucleus is described. The algorithm makes use of intensity information, edge magnitude information and both object and edge connectivity information. This automated procedure generates a closed contour precisely along the edge of the nucleus. The procedure was tested on a database of 3,680 red-green-blue images of thionin-SO2 and orange II-stained cervical cells obtained from normal and dysplastic samples. When used in conjunction with a simple threshold selection algorithm and an artifact removal routine, this edge relocation algorithm resulted in the correct segmentation of over 98% of the nuclei. Only 63 (1.7%) of all nuclei were incorrectly segmented.

Algorithms